Embryonic stem cells (ES cells) have multilineage potential and can differentiate into a variety of tissue cells. Therefore, a variety of studies have been carried out relating to the application of embryonic stem cells in the field of so-called regenerative medicine, in which cells lost due to disease, accidents, or the like are repaired and tissues are restored (for example, PTL 1).
ES cells have great diversity and can differentiate into a variety of cells. An example of a method for differentiating ES cells into a variety of cells is the formation of cell aggregates known as embryonic bodies (EB). These cell aggregates are formed by floating a culture of ES cells, iPS cells, and the like, and, when the ES cells are cultured for approximately two weeks in a state in which cell aggregates are formed, differentiation into a variety of cell types is observed. Therefore, the formation of embryonic bodies is used as one of the standard methods for investigating the pluripotent differentiation of cells.
Among the methods for culturing ES cells in a floating state, the most widely used method is hanging drop culture. Hanging drop culture is a method in which cells are cultured in culture fluids suspended in a water droplet shape. However, this method has problems of a low success rate of formation of embryonic bodies, incapability of microscopic observation, troublesome operations, and the like. In order to solve these problems, for example, culture vessels having water-insoluble cured membranes formed on the vessel inner surfaces by curing water-soluble resin membranes have been proposed (for example, PTL 2).
When clinical application is considered, research and development using human ES cells becomes necessary, but human ES cells have problems of a higher possibility of cell death and a greater difficulty in obtaining embryonic bodies compared with mouse ES cells. In order to solve these problems, for example, culture vessels have been proposed in which funnel shapes having an aperture angle in a range of 60 degrees to 100 degrees are provided as the well bottom portions and the center portions have concave roundness (for example, PTL 3). For research and development using human iPS cells as well, the same culture vessels have been proposed.
PTL 1 (Japanese Unexamined Patent Application, First Publication No. 2008-99662), PTL 2 (Japanese Unexamined Patent Application, First Publication No. 2008-178367), and PTL 3 (WO2013/183777) are incorporated into the present specification by reference.